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Microbiology and Biotechnology Letters

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Microbial Biotechnology (MB)  |  Cell Culture and Biomedical Engineering

Microbiol. Biotechnol. Lett.

Received: December 26, 2020; Revised: February 25, 2021; Accepted: February 25, 2021

Improving the Viability of Freeze-dried Probiotics Using a Lysine-based Rehydration Mixture

Karina Arellano1†, Haryung Park2†, Bobae Kim2, Subin Yeo2, Hyunjoo Jo3, Jin-Hak Kim4, Yosep Ji3, and Wilhelm H. Holzapfel1,2*

1Department of Advanced Convergence, 2HEM Inc., Business Incubator Center 103, Handong Global University, Pohang 37554, Republic of Korea 3HEM Inc., 404, Ace Gwanggyo Tower 3, Suwon 16229, Republic of Korea 4COSMAX NS Inc., Seongnam 13486, Republic of Korea

Correspondence to :
Wilhelm Holzapfel, 
wilhelm@woodapple.net

The probiotic market is constantly continuing to grow, concomitantly with a widening in the range and diversity of probiotic products. Probiotics are defined as live microorganisms that provide a benefit to the host when consumed at a proper dose; the viability of a probiotic is therefore of crucial importance for its efficacy. Many products undergo lyophilization for maintaining their shelf-life. Unfortunately, this procedure may damage the integrity of the cells due to stress conditions during both the freezing and (vacuum-) drying process, thereby impacting their functionality. We propose a lysine-based mixture for rehydration of freeze-dried probiotics for improving their viability during in vitro simulated gastric and duodenum stress conditions. Measurement of the zeta potential served as an indicator of cell integrity and efficacy of this mixture, while functionality was estimated by adhesion to a human enterocyte-like Caco-2 cell-line. The freeze-dried bacteria exhibited a significantly different zeta potential compared to fresh cultures; however, this condition could be restored by rehydration with the lysine mixture. Recovery of the surface charge was found to influence adhesion ability to the Caco-2 cell-line. The optimum lysine concentration of the formulation, designated “Zeta-bio”, was found to be 0.03 M for improving the viability of Lactiplantibacillus plantarum Lp-115 by up to 13.86% and a 7-strain mixture (400B) to 41.99% compared to the control rehydrated with distilled water. In addition, the lysine Zeta-bio formulation notably increased the adherence ability of lyophilized Lp-115 to the Caco-2 cell-line after subjected to the in vitro stress conditions of the simulated gastrointestinal tract passage.

Keywords: Zeta potential, probiotic, viability, cell adhesion, L-lysine

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